HGF STIMULATES STAT3 SYNTHESIS IN HUMAN RETINAL PIGMENT EPITHELIAL CELLS (hRPE)

Abstract

Abstract: : Purpose: The purpose of this study is to determine the effect of Tyrphostin AG490 (AG490), an inhibitor of STAT3 protein synthesis, on the hepatocyte growth factor (HGF) induced proliferation of human retinal pigment epithelial cells. Methods: Primary hRPE cell cultures were established from three human eyes. Cells were treated with AG490 in absence and presence of HGF. Cell proliferation was monitored by the trypan blue exclusion method and ³H–thymidine (³H–thy) incorporation. ¹⁴C–methionine labeled–intracellular signal transducer and activator of transcription–3 protein (¹⁴C–STAT3) synthesis was determined by immunoprecipitation and immunohistochemistry using STAT3 specific antibody. Data were analyzed by Student's 't' test. Results: HGF increased hRPE cell proliferation in a dose dependent manner. The addition of AG490 (50 µM) inhibited HGF stimulated hRPE cell proliferation as monitored by ³H–thy incorporation (11690.78±2327.89 vs. 3695.8±616.2, p<0.05, mean±SEM, n=3). HGF also increased ¹⁴C–STAT3 synthesis in a dose dependent manner. However, AG490 decreased ¹⁴C–STAT3 synthesis in a dose dependent manner (8600.30±1235.89 vs. 1713.72±125.08, p<0.05, mean±SEM, n=3). Immunohistochemical studies showed increased positive immunoreactivity of STAT3 protein in hRPE cells exposed of HGF. The addition of AG490 to wells containing HGF inhibited corresponding STAT3 protein immunoreactivity. Conclusions: AG490 inhibits HGF induced hRPE cell proliferation and intracellular STAT3 synthesis and therefore may be useful as a therapeutic agent to treat eye diseases involving abnormal hRPE cell proliferation.

Keywords: retinal pigment epithelium • growth factors/growth factor receptors • proliferation